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51. 发明授权

US07985575B2 Single protein production in living cells facilitated by a messenger RNA interferase 有权
标题翻译：由信使RNA干扰酶促进活细胞中的单一蛋白质生成
公开(公告)号：US07985575B2
公开(公告)日：2011-07-26
申请号：US11750314
申请日：2007-05-17
申请人： Masayori Inouye , Junjie Zhang , Motoo Suzuki
发明人： Masayori Inouye , Junjie Zhang , Motoo Suzuki
IPC分类号： C12N1/21
CPC分类号： C12N9/22 , C12N15/67 , C12N15/74
摘要： The present invention describes a single-protein production (SPP) system in living E. coli cells that exploits the unique properties of an mRNA interferase, for example, MazF, a bacterial toxin that is a single stranded RNA- and ACA-specific endoribonuclease, which efficiently and selectively degrades all cellular mRNAs in vivo, resulting in a precipitous drop in total protein synthesis. Concomitant expression of MazF and a target gene engineered to encode an ACA-less mRNA results in sustained and high-level (up to 90%) target expression in the virtual absence of background cellular protein synthesis. Remarkably, target synthesis continues for at least 4 days, indicating that cells retain transcriptional and translational competence despite their growth arrest. SPP technology works well for yeast and human proteins, even a bacterial integral membrane protein. This novel system enables unparalleled signal to noise ratios that should dramatically simplify structural and functional studies of previously intractable but biologically important proteins. The present invention also provides an optimized condensed single protein production system.
摘要翻译：本发明描述了利用mRNA干扰酶的独特性质的活的大肠杆菌细胞中的单蛋白质生产（SPP）系统，例如MazF，单链RNA-和ACA特异性内切核糖核酸酶的细菌毒素，其有效和选择性地降解体内所有细胞mRNA，导致总蛋白质合成的急剧下降。 MazF和旨在编码无ACA mRNA的靶基因的伴随表达导致在虚拟无背景细胞蛋白质合成中的持续和高水平（高达90％）的靶表达。值得注意的是，目标合成持续至少4天，表明细胞保留转录和翻译能力，尽管其生长停滞。 SPP技术适用于酵母和人类蛋白质，甚至细菌整合膜蛋白。这种新颖的系统能够实现无与伦比的信噪比，这将显着简化以前难以处理但生物重要的蛋白质的结构和功能研究。本发明还提供优化的冷凝单蛋白生产系统。

52. 发明申请

US20110097722A1 Cold Shock Protein Compositions and Methods and Kits for the Use Thereof 有权
标题翻译：冷冲击蛋白质组合物及其使用的方法和试剂盒
公开(公告)号：US20110097722A1
公开(公告)日：2011-04-28
申请号：US12919985
申请日：2009-03-02
申请人： Masayori Inouye , Sangita Phadtare , Ikunoshin Kato , Ling Zhu , Hiroyuki Mukai , Takashi Uemori , Kazue Nishiwaki
发明人： Masayori Inouye , Sangita Phadtare , Ikunoshin Kato , Ling Zhu , Hiroyuki Mukai , Takashi Uemori , Kazue Nishiwaki
IPC分类号： C12Q1/68 , C12N9/12 , C12P19/34
CPC分类号： C12Q1/6848 , C12Q2522/101
摘要： The present invention provides cold shock protein-containing compositions for improved DNA synthesis reactions with improved reactivity, methods for synthesizing DNA using such compositions, kits for use in such methods, and DNA compositions yielded by such methods. The present invention further provides cold shock protein-containing compositions for the identification of endoribonuclease cleavage sites, methods for identifying endoribonuclease cleavage sites using such compositions, and kits for use in such methods.
摘要翻译：本发明提供了用于改善反应性的DNA合成反应的改善的含有冷休克蛋白的组合物，使用这种组合物合成DNA的方法，用于此类方法的试剂盒以及通过这些方法产生的DNA组合物。本发明进一步提供用于鉴定内切核糖核酸酶切割位点的含有冷休克蛋白的组合物，使用这种组合物鉴定内切核糖核酸酶切割位点的方法，以及用于此类方法的试剂盒。

53. 发明申请

US20090047742A1 Inhibition of mRNA Interferase-Induced Apoptosis in BAK-Deficient and BAK- and Bax-Deficient Mammalian Cells 审中-公开
标题翻译：抑制BAK缺陷型和BAK-和Bax-缺陷型哺乳动物细胞中mRNA干扰素诱导的细胞凋亡
公开(公告)号：US20090047742A1
公开(公告)日：2009-02-19
申请号：US12064070
申请日：2006-08-22
申请人： Tsutomu Shimazu , Kurt Degenhardt , Eileen White , Masayori Inouye
发明人： Tsutomu Shimazu , Kurt Degenhardt , Eileen White , Masayori Inouye
IPC分类号： C12N15/87 , C12N5/06
CPC分类号： A61K48/0066 , A61K48/00 , C07K14/4747 , C12N9/22 , C12N2800/40
摘要： Ribonucleases, antibiotics, bacterial toxins and viruses inhibit protein synthesis, which results in apoptosis in mammalian cells. How the BCL-2 family of proteins regulates apoptosis in response to shutoff of protein synthesis is not known. According to the present invention, an Escherichia coli toxin MazF inhibited protein synthesis by cleavage of cellular mRNA, and induced apoptosis in mammalian cells. MazF-induced apoptosis required proapoptotic BAK and its upstream regulator, the proapoptotic BH3-only protein NBK/BIK, but not BIM, PUMA or NOXA. Furthermore, NBK/BIK- or BAK-deficient cells were resistant to cell death induced by pharmacologic inhibition of translation and by virus-mediated shutoff of protein synthesis. Thus, the BH3-only protein NBK/BIK is the apical regulator of a BAK-dependent apoptotic pathway in response to shutoff of protein synthesis. Although NBK/BIK is dispensable for development, it is the BH3-only protein targeted for inactivation by viruses, suggesting that it plays a role in pathogen/toxin response through apoptosis activation.
摘要翻译：核糖核酸酶，抗生素，细菌毒素和病毒抑制蛋白质合成，导致哺乳动物细胞凋亡。 BCL-2家族蛋白质如何调节蛋白质合成关闭反应的凋亡是未知的。根据本发明，大肠杆菌毒素MazF通过切割细胞mRNA抑制蛋白质合成，并诱导哺乳动物细胞的细胞凋亡。 MazF诱导的细胞凋亡需要促凋亡BAK及其上游调节因子，即促凋亡BH3蛋白NBK / BIK，而不是BIM，PUMA或NOXA。此外，NBK / BIK-或BAK缺陷细胞对由翻译的药理学抑制和蛋白质合成的病毒介导的切断引起的细胞死亡具有抗性。因此，仅BH3蛋白NBK / BIK是响应于蛋白质合成关闭的BAK依赖性凋亡途径的顶端调节剂。虽然NBK / BIK是不可开发的，但是仅针对通过病毒灭活的BH3蛋白，这表明它通过凋亡激活在病原体/毒素反应中起作用。

54. 发明申请

US20060054941A1 Multifunctional biosensor based on ZnO nanostructures 审中-公开
标题翻译：基于ZnO纳米结构的多功能生物传感器
公开(公告)号：US20060054941A1
公开(公告)日：2006-03-16
申请号：US11119475
申请日：2005-04-29
申请人： Yicheng Lu , Zheng Zhang , Nuri Emanetoglu , Masayori Inouye , Oleg Mirochnitchenko
发明人： Yicheng Lu , Zheng Zhang , Nuri Emanetoglu , Masayori Inouye , Oleg Mirochnitchenko
IPC分类号： C12Q1/68 , H01L29/82
CPC分类号： G01N21/6428 , C12Q1/6825 , C12Q1/6837 , G01N27/4145 , G01N27/4146 , G01N29/022 , G01N33/54373 , G01N33/5438 , G01N2021/6432 , G01N2291/011 , G01N2291/014 , G01N2291/0255 , G01N2291/0256 , G01N2291/0423 , G01N2291/0426 , C12Q2565/501 , C12Q2565/607
摘要： The present invention provides the multifunctional biological and biochemical sensor technology based on ZnO nanostructures. The ZnO nanotips serve as strong DNA or protein molecule binding sites to enhance the immobilization. Patterned ZnO nanotips are used to provide conductivity-based biosensors. Patterned ZnO nanotips are also used as the gate for field-effect transistor (FET) type sensors. Patterned ZnO nanotips are integrated with SAW or BAW based biosensors. These ZnO nanotip based devices operate in multimodal operation combining electrical, acoustic and optical sensing mechanisms. The multifunctional biosensors can be arrayed and combined into one biochip, which will enhance the sensitivity and accuracy of biological and biochemical detection due to strong immobilization and multimodal operation capability. Such biological and biochemical sensor technology are useful in detection of RNA-DNA, DNA-DNA, protein-protein, protein-DNA and protein-small molecules interaction. It can be further applied for drug discovery, and for environmental monitoring and protection.
摘要翻译：本发明提供了基于ZnO纳米结构的多功能生物和生物化学传感器技术。 ZnO纳米尖端作为强的DNA或蛋白质分子结合位点来增强固定。图案化的ZnO纳米尖端用于提供基于导电性的生物传感器。图案化的ZnO纳米技术也用作场效应晶体管（FET）型传感器的栅极。图案化的ZnO纳米片与SAW或基于BAW的生物传感器集成。这些基于ZnO纳米管的器件在电气，声学和光学感测机构的多模式操作中工作。多功能生物传感器可以排列并组合成一个生物芯片，由于强固定和多模态操作能力，提高了生物和生化检测的灵敏度和准确性。这种生物和生化传感器技术可用于RNA-DNA，DNA-DNA，蛋白质 - 蛋白质，蛋白质 - DNA和蛋白质 - 小分子相互作用的检测。可进一步应用于药物发现，环境监测和保护。

55. 发明申请

US20050272924A1 Cold shock inducible expression and production of heterologous polypeptides 失效
标题翻译：冷休克诱导表达和产生异源多肽
公开(公告)号：US20050272924A1
公开(公告)日：2005-12-08
申请号：US10506192
申请日：2003-02-25
申请人： Masayori Inouye , Sangita Phadtare , Bing Xia , Guoliang Qing , Haiping Ke
发明人： Masayori Inouye , Sangita Phadtare , Bing Xia , Guoliang Qing , Haiping Ke
IPC分类号： C12N15/09 , C07H21/04 , C07K14/47 , C12N1/15 , C12N1/21 , C12N15/63 , C12N15/67 , C12N15/70 , C12P21/02 , C12P21/06 , C12Q1/00
CPC分类号： C12N15/67 , C07K2299/00 , C12N15/70 , Y10T436/24
摘要： The present invention relates to a DNA molecule or vector and a host cell containing the DNA molecule or vector which can be used to produce a heterologous polypeptide under conditions that elicit a cold shock response in the host cell. The DNA molecule and vector include a nucleotide sequence encoding a heterologous polypeptide and a promoter and 5′-UTR from a cold shock inducible gene which directs its expression. IN addition, an AT-rich sequence that enhances translation under cold shock inducible conditions is either present in the coding sequence of the heterologous polypeptide or in an additional element inserted between the coding sequence and the cold shock inducible promoter and 5′-UTR.
摘要翻译：本发明涉及DNA分子或载体以及含有DNA分子或载体的宿主细胞，其可以在宿主细胞中引起冷休克反应的条件下用于产生异源多肽。 DNA分子和载体包括编码异源多肽的核苷酸序列和引导其表达的冷休克诱导型基因的启动子和5'-UTR。此外，在冷休克诱导条件下增强翻译的富含AT的序列存在于异源多肽的编码序列中，或存在于插入编码序列和冷休克诱导型启动子和5'-UTR之间的另外的元件中。

56. 发明授权

US06610533B1 Cold-shock regulatory elements, constructs thereof, and methods of use 失效
标题翻译：冷休克调节元件，其构建体和使用方法
公开(公告)号：US06610533B1
公开(公告)日：2003-08-26
申请号：US09516667
申请日：2000-03-01
申请人： Masayori Inouye , Nan Wang , Kunitoshi Yamanaka
发明人： Masayori Inouye , Nan Wang , Kunitoshi Yamanaka
IPC分类号： C12N1500
CPC分类号： C07K14/245
摘要： A fourth cold shock protein of the Escherichia coli CspA family is disclosed, as are the regulatory elements of the 5′ UTR of the corresponding gene. The cspI gene is located at 35.2 min on the E. coli chromosome map, and CspI shows 70, 70, and 79% identity of CspA, CspB, and CspG, respectively. The 5′-untranslated region of the cspI mRNA consists of 145 bases and causes a negative effect on cspI expression at 37° C. The cspI mRNA was very unstable at 37° C. but was stabilized upon cold shock. The 5′ UTR of cspI can enhance the translation of cold shock inducible genes under conditions that elicit a cold shock response in bacteria.
摘要翻译：公开了大肠杆菌CspA家族的第四种冷休克蛋白，以及相应基因的5'UTR的调节元件。 cspI基因位于大肠杆菌染色体图上的35.2分钟，CspI分别显示CspA，CspB和CspG的70,70和79％同一性。 cspI mRNA的5'非翻译区由145个碱基构成，并在37℃引起cspI表达的负面影响.cspI mRNA在37℃非常不稳定，但在冷休克时稳定。 cspI的5'UTR可以在引起细菌中的冷休克反应的条件下增强冷休克诱导型基因的翻译。

57. 发明授权

US5719021A Protein activation 失效
标题翻译：蛋白激活
公开(公告)号：US5719021A
公开(公告)日：1998-02-17
申请号：US923260
申请日：1992-07-31
申请人： Masayori Inouye
发明人： Masayori Inouye
IPC分类号： C12N9/00 , C07K1/00 , C07K1/113 , C07K14/575 , C07K14/62 , C07K14/745 , C07K14/76 , C12N9/56 , C12P21/00 , C12P21/02 , C12Q1/68 , C07K14/435 , C12N9/96
CPC分类号： C07K1/1133 , Y10S435/839
摘要： A method is disclosed for producing a biochemically active polypeptide from a biochemically inactive polypeptide. The polypeptide is normally but need not be expressed in a precursor form containing a pro-sequence. The inactive polypeptide is reacted with a tailor-made activating peptide. The activating peptide can be synthetic or made by recombinant DNA procedure. The activating peptide is a peptide which contains one or more functional domains which are necessary for folding the inactive polypeptide into a biochemically active conformation. The activating peptide may but need not contain a sequence of amino acids which is identical to the sequence of the natural occurring pro-sequence of the polypeptide. Also, a method is disclosed which permits to identify the one or more functional domains in the pro-sequence of a polypeptide which contribute(s) to the folding of the inactive polypeptide into a biochemically active conformation. The invention relates also to a tailor-made activating peptide (synthetic or by recombinant DNA) and to the biochemically active polypeptide. The protein activation method and the biochemically active proteins are of major utility in their broad applications.
摘要翻译：公开了用于从生物化学惰性多肽生产生物化学活性多肽的方法。多肽通常但不需要以含有前序列的前体形式表达。无活性多肽与定制的活化肽反应。活化肽可以是合成的或通过重组DNA程序制备的。活化肽是含有一个或多个将无活性多肽折叠成生物化学活性构象所必需的功能结构域的肽。活化肽可以不必含有与多肽的天然存在的前序序列相同的氨基酸序列。此外，公开了一种方法，其允许鉴定有助于将无活性多肽折叠成生物化学活性构象的多肽的前序序列中的一个或多个功能结构域。本发明还涉及定制的活化肽（合成的或通过重组DNA）和生物化学活性多肽。蛋白激活方法和生物化学活性蛋白质在广泛应用中具有重要的实用价值。

58. 发明授权

US5714575A Nucleic acids sequence, stress-induced proteins and uses thereof 失效
标题翻译：核酸序列，应激诱导的蛋白质及其用途
公开(公告)号：US5714575A
公开(公告)日：1998-02-03
申请号：US203806
申请日：1994-03-01
申请人： Masayori Inouye , Pamela Jones , Jean-Pierre Etchegaray , Weining Jiang , N. Stephen Pollitt , Joel Goldstein
发明人： Masayori Inouye , Pamela Jones , Jean-Pierre Etchegaray , Weining Jiang , N. Stephen Pollitt , Joel Goldstein
IPC分类号： C07K14/245 , C12N15/70 , C12N15/82 , C07K14/24
CPC分类号： C12N15/70 , C07K14/245 , C12N15/8273
摘要： A family of stimuli-induced in particular stress or cold-shock induced genes and proteins are disclosed which have conserved amino acid domains. Nucleic acid sequences of the genes and the promoters are also described. Various utilities of the promoters and of the proteins are disclosed.
摘要翻译：公开了一种具有特定应激或冷休克诱导的基因和蛋白质的刺激物，其具有保守的氨基酸结构域。还描述了基因和启动子的核酸序列。公开了启动子和蛋白质的各种效用。

59. 发明授权

US5714323A Over expression of single-stranded molecules 失效
标题翻译：单链分子的过表达
公开(公告)号：US5714323A
公开(公告)日：1998-02-03
申请号：US318867
申请日：1995-05-04
申请人： Atushi Ohshima , Sumiko Inouye , Masayori Inouye
发明人： Atushi Ohshima , Sumiko Inouye , Masayori Inouye
IPC分类号： C12N15/10 , C12N15/70 , C12Q1/68 , C12P19/34
CPC分类号： C12N15/70 , C12N15/10 , C12N2310/111
摘要： A method of synthesis of new and useful single-stranded DNAs which have a stem-loop configuration (ss-slDNA). The method is an in vivo or an in vitro synthesis. Replicating vehicles which produce these ss-slDNAs. The ss-slDNAs are described. Uses for these slDNAs are disclosed. They can be used for introducing random mutations, they lend themselves for replication by a variant of the PCR method. They can also be used for regulating gene function. Other uses are disclosed.
摘要翻译： PCT No.PCT / US94 / 02169 Sec。 371日期：1995年5月4日 102（e）日期1995年5月4日PCT 1994年3月1日PCT公布。出版物WO94 / 20639 日期1994年9月15日一种合成具有茎 - 环构型（ss-slDNA）的新型和有用的单链DNA的方法。该方法是体内或体外合成。复制产生这些ss-SLDNA的载体。描述了ss-SLDNA。公开了这些用于这些slDNA的用途。它们可用于引入随机突变，它们通过PCR方法的变体使其复制。它们也可用于调节基因功能。披露了其他用途。

60. 发明授权

US5405775A Retrons coding for hybrid DNA/RNA molecules 失效
标题翻译：编码杂交DNA / RNA分子的核苷酸
公开(公告)号：US5405775A
公开(公告)日：1995-04-11
申请号：US518749
申请日：1990-05-02
申请人： Masayori Inouye , Sumiko Inouye
发明人： Masayori Inouye , Sumiko Inouye
IPC分类号： C12N9/12 , C12N15/10 , C12P19/34 , C12N1/21 , C12N15/52 , C12N15/70
CPC分类号： C12N9/1276 , C12N15/10 , C12P19/34
摘要： A multicopy single-stranded DNA (msDNA) synthesizing system in E. coli is disclosed. The use of the msDNA system to synthesize cDNA in vivo is disclosed. Construction of synthetic msDNA is also disclosed. Also processes for gene amplification and for producing a stable RNA are disclosed.
摘要翻译：公开了在大肠杆菌中的多拷贝单链DNA（msDNA）合成系统。公开了使用msDNA系统在体内合成cDNA。还公开了合成msDNA的构建。还公开了用于基因扩增和产生稳定的RNA的过程。

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